32份木薯种质对疫霉根腐病的抗性评价和农艺性状分析

通过田间自然发病和室内离体接种方法对32份木薯种质进行了棕榈疫霉根腐病抗病性评价,结果表明,供试的32份木薯种质中,高抗种质7份,抗病种质7份,中感种质8份,感病种质7份,高感种质3份。其中高抗种质为‘H360’、‘华南11号’、‘华南8号’、‘H588’、‘桂热3号’、‘H873’和‘H971’;抗病种质为:‘F556’、‘H502’、‘GR911’、‘C-4’、‘F10’、‘南植188’和‘H47’。对其中14份抗棕榈疫霉根腐病木薯种质的农艺性状进行了鉴定和评价,结果表明‘华南8号’、‘GR911’、‘F556’、‘C-4’、‘H360’可以进一步加以利用。     

小麦赤霉病菌对叶菌唑的抗性风险分析

采用菌丝生长速率法,测定了2012–2014年采自我国江苏、安徽、山东和河南4个省份的100株小麦赤霉病菌对叶菌唑的敏感性,并通过室内药剂驯化获得叶菌唑抗性突变体,研究了抗性突变体的适合度及CYP51基因序列和表达量.结果表明:叶菌唑对供试菌株的EC50值范围为0.04～0.51μg/mL,平均EC50值为(0.18±...     

水稻稻瘟病菌对烯肟菌胺的抗性风险评估及抗性机制初探

 采用菌丝生长速率法测定了100株采自我国主要水稻产区的水稻稻瘟病菌对烯肟菌胺的敏感性, 结果表明, 其EC₅₀分布于0.011 1~0.295 6 μg·mL^-1, 平均EC₅₀=(0.078 6±0.056 1) μg·mL^-1。供试菌株对烯肟菌胺的敏感性分布呈单侧峰曲线, 未出现抗药性亚群体, 可将该曲线作为稻病瘟菌对烯肟菌胺的敏感性基线。通过室内药剂驯化获得了7株抗药突变体, 突变频率为1.11×10^-4, 其中2株高抗突变体NJ0811-I和A10的抗性水平大于1 000倍, 抗药性性状能稳定遗传, 致病力显著弱于其亲本菌株;5株低抗突变体抗性水平在2.05～4.55倍之间, 抗药稳定性差, 适合度与亲本无显著性差异。交互抗药性结果表明, 烯肟菌胺与嘧菌酯存在正交互抗药性, 与田间防治稻瘟病常用药剂稻瘟灵、异稻瘟净无交互抗药性。综合分析表明, 稻瘟病菌对烯肟菌胺可能存在低到中等抗性风险。进一步克隆了抗药突变体及其亲本的cytb基因, CYTB氨基酸序列比对结果表明, 2株高抗突变体均在143位由甘氨酸突变为丝氨酸(G143S), 建立了高抗菌株的AS-PCR分子检测方法;而5株低抗突变体cytb基因未发生点突变, 推测可能存在其他的抗性分子机制。     

禾谷镰刀菌对苯基吡咯类杀菌剂咯菌腈的抗性机制

由禾谷镰刀菌引起的小麦赤霉病是世界小麦生产上的重要真菌病害。为了进一步明确禾谷镰刀菌对苯基吡咯类杀菌剂咯菌腈产生抗性的机制,本文以前期室内通过药剂驯化方式得到的4株禾谷镰刀菌对咯菌腈的高水平抗性突变体(其抗性倍数在318.2～782.9之间)为主要研究材料,采用生物测定及分子生物学等方法开展了禾谷镰刀菌对咯菌腈的抗性机制研究。结果表明：供试禾谷镰刀菌抗咯菌腈突变体对小麦幼穗的致病力降低了约50%,部分菌株(2XZ-4R)甚至完全丧失了对小麦的致病能力;抗性突变体对渗透胁迫(0.5 mol/L NaCl,1.0 mol/L MgCl₂, 1.0 mol/L葡萄糖或1.0 mol/L甘露醇)高度敏感,且菌丝生长抑制率较敏感菌株降低约50%以上,表明其环境适合度显著下降。同时,抗性突变体中苯丙氨酸解氨酶(PAL)、过氧化物酶(POD)和多酚氧化酶(PPO)活性较敏感菌株均升高2倍以上。分子生物学分析发现,供试抗性突变体中候选靶标基因(FgOs1和FgOs5)的表达量显著下调(P<0.05),推测FgOs1和FgOs5可能参与了禾谷镰刀菌对咯菌腈抗性的形成过程。总之...     

Characterization of Laboratory Mutants of Botrytis cinerea Resistant to QoI Fungicides

Mutants of Botrytis cinerea with moderate and high resistance to pyraclostrobin, a Qo inhibitor of mitochondrial electron transport at the cytochrome bc ₁ complex, were isolated at a high mutation frequency, after nitrosoguanidine mutagenesis and selection on medium containing pyraclostrobin and salicylhydroxamate (SHAM), a specific inhibitor of cyanide-resistant (alternative) respiration. Oxygen uptake in whole cells was strongly inhibited in the wild-type strain by pyraclostrobin and SHAM, but not in the mutant isolates. Cross-resistance studies with other Qo and Qi inhibitors (QoIs and QiIs) of cytochrome bc ₁ complex of mitochondrial respiration showed that the mutation(s) for resistance to pyraclostrobin also reduced the sensitivity of mutant strains to other QoIs as azoxystrobin, fluoxastrobin, trifloxystrobin and picoxystrobin, but not to famoxadone and to the QiIs cyazofamid and antimycin-A. An increased sensitivity of pyraclostrobin-resistant strains to the carboxamide boscalid, an inhibitor of complex II, and to the anilinopyrimidine cyprodinil, a methionine biosynthesis inhibitor, was observed. Moreover, no effect of pyraclostrobin resistance mutation(s) on fungitoxicity of the hydroxyanilide fenhexamid, the phenylpyrrole fludioxonil, the benzimidazole benomyl, and to the phenylpyridinamine fluazinam, which affect other cellular pathways, was observed. Study of fitness parameters in the wild-type and pyraclostrobin-resistant mutants of B. cinerea showed that most mutants had a significant reduction in the sporulation, conidial germination and sclerotia production. Experiments on the stability of the pyraclostrobin-resistant phenotype showed a reduction of resistance, mainly in moderate resistant strains, when the mutants were grown on inhibitor-free medium. However, a rapid recovery of the resistance level was observed after the mutants were returned to a selective medium. Studies on the competitive ability of mutant isolates against the wild-type parent strain, by applications of a mixed conidial population, showed that, in vitro, all mutants were less competitive than the wild-type strain. However, the competitive ability of high resistant mutants was higher than the moderate ones. Pathogenicity tests on cucumber seedlings showed that all mutant strains tested exhibited an infection ability similar with the wild-type parent strain. Preventive applications of the commercial product of F-500 25EC (pyraclostrobin) were effective against lesion development on cotyledons by the wild-type, but ineffective, even at high concentrations, against disease caused by the pyraclostrobin-resistant isolates. Boscalid (F-510 50WG) was found equally effective against the disease caused by the wild-type or pyraclostrobin-resistant mutants. This is the first report indicating the appearance of B. cinerea strains resistant to QoI fungicides by the biochemical mechanism of site modification and the risk for field resistance.     

小麦赤霉病菌对多菌灵的抗药性研究

测定了1976 年从江苏南京采集的54 株小麦赤霉病菌菌株对多菌灵的敏感性,EC50值在0. 2617～ 0. 6544 μg. mL - 1 之间, 1983 年在同一地点采集的76 株, EC50 值相应为0. 2517～ 0. 7050 μg. mL- 1 , 而1998 年从浙江、湖北、上海、福建、安徽、江苏各地采集的104 株菌株EC50值为0. 2478～ 6. 4574 Lg. mL- 1 , 表明湖北、上海、浙江等地田间已检测到小麦赤霉病菌抗多菌灵菌株。紫外光诱导分生孢子也获得了该病菌抗多菌灵突变株, 其EC50 值为14. 1993 Lg. mL - 1 。抗药性突变体JPR 与敏感亲本菌株JPS 相比, 在菌丝生长、产孢量方面无明显差异, 但JPR 孢子萌发率为57. 1% , 而JPS 为100% , 而且50% 孢子萌发的时间较野生亲本菌株滞后12 h。JPR 产生脱氧雪腐镰刀菌烯醇(DON 毒素) 3. 90 μg mL - 1 , 而JPS 产生的DON 毒素为9. 28μg. mL - 1 。     

西瓜蔓枯病菌啶酰菌胺抗性突变体的生物学特性研究及其Sdh B基因位点检测

为评价西瓜蔓枯病菌对啶酰菌胺的抗性风险,了解其抗性机理,室内通过药剂驯化方法获得2株啶酰菌胺的抗性突变体XF21-3和YC60-1,测定了抗性突变体的生物学特性,并通过对Sdh B基因片段的测序比对,分析了西瓜蔓枯病菌对啶酰菌胺的抗性机理。生物测定结果表明:啶酰菌胺对2株抗性突变体的EC50值分别为108和124 μg/mL,抗性倍数（RR）分别为1 007和1 347,均为高抗菌株;抗性突变体的菌丝生长速率和产孢量均大于亲本菌株,但其致病性与亲本菌株无显著差异,对外界环境渗透压的敏感性低于亲本菌株;此外,啶酰菌胺与萎锈灵、戊唑醇、乙霉威及醚菌酯之间均不存在交互抗性,但与噻呋酰胺之间存在交互抗性。Sdh B基因片段测序及比对结果表明,高抗性突变体中Sdh B亚基277位上的氨基酸所对应的碱基由CAC突变为TAC,即由组氨酸（His）突变为酪氨酸（Tyr）。研究表明,西瓜蔓枯病菌在药剂选择压力下容易形成啶酰菌胺的抗性群体,且抗性突变体的离体适合度高于亲本菌株,此外,啶酰菌胺与同类型杀菌剂噻呋酰胺之间存在交互抗性,因此认为西瓜蔓枯病菌对啶酰菌胺具有中等抗性风险;同时进一步验证了Sdh B亚基277位上的氨基酸突变（His→Tyr,CAC→TAC）是西瓜蔓枯病菌对啶酰菌胺产生抗性的原因。     

水稻白叶枯病菌室内链霉素抗性菌株的诱导及其抗性分子机制和风险评价

通过紫外光照射诱导获得了6株水稻白叶枯病菌Xanthomonas oryzae pv.oryzae抗链霉素突变体。测得链霉素对水稻白叶枯病菌敏感菌株ZJ173及其抗性突变体的最低抑制浓度(MIC)分别为0.10和600 μg/mL;对敏感菌株的有效抑制中浓度(EC50)为0.03 μg/mL,对抗性菌株的平均EC50值为11.64 μg/mL,平均抗性倍数为388。通过PCR扩增了敏感菌株ZJ173及5株抗性菌株的rpsL基因(编码S12核糖体蛋白)和rrs基因(编码16S rRNA),并检测了strA基因是否存在。序列分析表明,5株被测抗性菌株的rpsL基因均发生了突变,其中4株在氨基酸43位、1株在88位,均由赖氨酸突变为精氨酸,而rrs基因未发生突变,strA基因未被检测到。表明实验室诱导获得的水稻白叶枯病菌抗性菌株对链霉素的抗药性是由rpsL基因突变引起的。抗性风险研究表明,抗性突变体的抗药性在无药剂压力下可稳定保持,其致病性、生长速率与敏感菌株相比无明显差异,竞争性低于或略低于敏感菌株,抗性自发突变率较高,且抗性突变为单一位点突变,病害循环为多循环,因此由rpsL基因突变引起的水稻白叶枯病菌对链霉素的抗性风险较高。     

Characterization of fludioxonil-resistant and pyrimethanil-resistant phenotypes of Penicillium expansum from apple

Penicillium expansum is the primary cause of blue mold, a major postharvest disease of apple. Fludioxonil and pyrimethanil are two newly registered postharvest fungicides for pome fruit in the United States. To evaluate the potential risk of resistance development in P. expansum to the new postharvest fungicides, one isolate of each of thiabendazole-resistant (TBZ-R) and -sensitive (TBZ-S) P. expansum was exposed to UV radiation to generate fungicide-resistant mutants. Four fludioxonil highly-resistant mutants (EC(50) > 1,000 microg/ml) and four pyrimethanil-resistant mutants (EC(50) > 10 microg/ml) were tested for sensitivities to thiabendazole, fludioxonil, and pyrimethanil, and fitness parameters including mycelial growth, sporulation on potato dextrose agar (PDA), sensitivity to osmotic stress, and pathogenicity and sporulation on apple fruit. The stability of resistance of the mutants was tested on PDA and apple fruit. Efficacy of the three fungicides to control blue mold incited by the mutants was evaluated on apple fruit. Six fungicide-resistant phenotypes were identified among the parental wild-type isolates and their mutants based upon their resistance levels. All four fludioxonil highly-resistant mutants were sensitive to pyrimethanil and retained the same phenotypes of resistance to TBZ as the parental isolates. All four pyrimethanil-resistant mutants had a low level of resistance to fludioxonil with a resistance factor >15. The two pyrimethanil-resistant mutants derived from a TBZ-S isolate became resistant to TBZ at 5 microg/ml. After 20 successive generations on PDA and four generations on apple fruit, the mutants retained the same phenotypes as the original generations. All mutants were pathogenic on apple fruit at both 0 and 20 degrees C, but fludioxonil highly-resistant mutants were less virulent and produced fewer conidia on apple fruit than pyrimethanil-resistant mutants and their parental wild-type isolates. Compared with the parental isolates, all four fludioxonil highly-resistant mutants had an increased sensitivity to osmotic stress on PDA amended with NaCl, while the pyrimethanil-resistant mutants did not. Pyrimethanil was effective against blue mold caused by fludioxonil-resistant mutants at both 0 and 20 degrees C. Pyrimethanil and fludioxonil reduced blue mold incited by pyrimethanil-resistant mutants during 12-week storage at 0 degrees C but were not effective at 20 degrees C. TBZ was not effective against pyrimethanil-resistant mutants derived from TBZ-S wild-type isolates at room temperature but provided some control at 0 degrees C. The results indicate that: (i) a fitness cost was associated with fludioxonil highly resistant mutants of P. expansum in both saprophytic and pathogenic phases of the pathogen but not pyrimethanil-resistant mutants; (ii) pyrimethanil possessed a higher risk than fludioxonil in the development of resistance in P. expansum; and (iii) triple resistance to the three apple-postharvest fungicides could emerge and become a practical problem if resistance to pyrimethanil develops in P. expansum populations.     

陕西省小麦赤霉病菌对多菌灵敏感性研究

 The mycelium growth rate method was used to test the sensitivity to carbendazim (MBC) at distinctive concentrations in 136 isolates of Fusarium graminearum from 19 counties of 6 districts in Shaanxi Province in 2008. The distinctive MBC concentration was 4 mg / L for testing of resistance and sensitivity. The results showed that average 50% effective concentration (EC50 ) of 136 tested sensitive isolates were (0. 908 6 ± 0. 062 3) mg / L. All the isolates were sensitive to MBC. The fungicide of MBC could be continually applied wheat production in Shaanxi.     

Impact of fludioxonil resistance on fitness and cross-resistance profiles of Altrenaria solani laboratory mutants

Sensitivity and inherent resistance risk of Alternaria solani to fludioxonil, cross-resistance profiles and the potential implications of resistance mutations on fitness parameters were investigated. Fludioxonil was highly effective against a wild type A. solani field strain both in vitro (EC₅₀?=?0.05 μg/mL) and in preventive applications on artificially inoculated tomato fruit. Mutants with low [Resistance factor (Rf): 15 based on EC₅₀], medium (Rf: 150–300) and high (Rf: > 1000) levels of phenylpyrrole resistance were isolated from the wild type strain at high frequencies following mutagenesis with UV irradiation and selection on fludioxonil containing medium. Resistant isolates retained their resistance levels even after 9 subcultures on fungicide-free growth medium while they could express their resistant phenotypes in planta. Investigation of cross-resistance relationships showed that fludioxonil resistance mutations also reduce the sensitivity of mutant strains to the aromatic hydrocarbon fungicide quintozene as well as the dicarboximides iprodione and vinclozolin. No cross-resistance was observed between fludioxonil and fungicides with different modes of action such as the sterol biosynthesis inhibitors (DMIs) imazalil and flusilazole and the carboxamide boscalid. All fludioxonil resistant isolates were more sensitive to the anilinopyrimidine pyrimethanil, while only two isolates were less sensitive to the QoI pyraclostrobin compared to the wild-type strain. Study of fitness determining parameters showed that resistance mutation(s) had no adverse effects on mycelial growth, conidial germination and sensitivity to osmotic stress while they had a pleiotropic effect on virulence and conidia production in resistant mutants. Results of the present study indicate that fludioxonil is a highly effective fungicide against A. solani, while the risk of resistance development to this fungicide is considered to be medium making fludioxonil an ideal alternative to high risk fungicides such as boscalid and pyraclostrobin whose performance against early blight has already been compromised by resistance development.

     

PIRA-PCR for detection of Fusarium graminearum genotypes with moderate resistance to carbendazim

PIRA-PCR ( p rimer- i ntroduced r estriction a nalysis PCR) was developed to detect isolates of Fusarium graminearum with moderate resistance to carbendazim, a methyl benzimidazole carbamate (MBC)-group fungicide. Two primer pairs were designed and synthesized according to the nucleotide sequence of the β ₂-tubulin gene from F. graminearum. Fragments of 164 bp were amplified by nested PCR from isolates differing in carbendazim sensitivity. A Hin dIII restriction enzyme recognition site was introduced artificially by inner primers to detect a mutation at codon 167, and Taa I ( Tsp 4CI) restriction enzyme was used to detect a mutation at codon 200. The sensitivity of isolates to carbendazim was determined by analyzing electrophoresis patterns of the resulting PCR products after simultaneous digestion with both Hin dIII and Taa I. Results from PIRA-PCR and a conventional method (mycelial growth on agar) were identical but PIRA-PCR required only 7–8 h while the conventional method required 5–7 days. This study demonstrates that PIRA-PCR not only monitors the appearance of moderately resistant isolates, but can be useful for detecting genotypes of F. graminearum with moderate resistance to carbendazim.     

黄瓜霜霉病菌对双炔酰菌胺的敏感基线及其抗性突变体生物学性状研究

为建立黄瓜霜霉病菌对双炔酰菌胺的敏感基线及评估其抗性风险,采用叶盘漂浮法测定了采自河北、山东未使用过羧酸酰胺类(CAAs)药剂地区的69株黄瓜霜霉病菌(Pseudoperonospora cubensis)对双炔酰菌胺的敏感性,并对黄瓜霜霉病菌抗双炔酰菌胺突变体的获得方法及突变体的生物学性状进行了研究。结果表明:69株黄瓜霜霉病菌对双炔酰菌胺的平均EC50值为(0.358±0.144)μg/mL,不同敏感性菌株的频率呈连续单峰曲线分布,未发现敏感性下降的亚群体,因此可将其作为黄瓜霜霉病菌对双炔酰菌胺的敏感基线;通过药剂驯化的方法未获得黄瓜霜霉病菌抗双炔酰菌胺的突变体;而通过紫外诱导的方法获得了6个抗双炔酰菌胺的突变体,其抗性水平介于5.74~22.96倍之间,突变频率为1.09×10-7,适合度显著低于其亲本菌株,且抗药性不能稳定遗传;双炔酰菌胺与甲霜灵、嘧菌酯、霜脲氰、氟吡菌胺之间无交互抗性关系,与烯酰吗啉之间有交互抗性关系。据此推测黄瓜霜霉病菌对双炔酰菌胺的抗性风险为低到中等。     

致病疫霉对缬菌胺抗性突变体生物学特性及抗性风险量化评估

 通过研究致病疫霉对缬菌胺抗性突变体的生物学特性,评估致病疫霉对缬菌胺产生抗性的风险。采用菌丝生长速率法检测田间菌株对缬菌胺的敏感性;通过比较抗性突变体在无药条件下继代培养前后对缬菌胺的敏感性来确定其抗性稳定性;采用离体叶片法测定抗性突变体及其亲本敏感菌株的适合度和竞争力;通过量化7个基本抗性因素的风险值评估致病疫霉对缬菌胺的基本抗性风险。结果表明:在2018年从内蒙、河北、贵州、四川、黑龙江等五省(自治区)采集分离的153个菌株和2019年从河北、内蒙采集分离的40个菌株中低抗菌株分别占31.4%和35.0%,敏感菌株分别占68.6%和65.0%,抗性指数均为0.34,抗性倍数分别为1.5和2.1,未发现中抗和高抗菌株;8个抗性突变体在无药条件下继代培养10代后,少数抗性突变体的抗药性不能稳定遗传;6个抗性突变体的适合度均显著低于其亲本敏感菌株;4个抗性突变体与其亲本菌株的孢子囊不同比率混合物分别在离体叶片上继代培养1、3、7代的抗性频率显著低于初始抗性频率或与初始抗性频率无显著差异;致病疫霉对缬菌胺7个基本抗性因素的风险值之和为9,推测致病疫霉对缬菌胺的基本抗性风险为低至中等。建议加强致病疫霉菌群体对缬菌胺的抗性监测,将缬菌胺与不同作用机理的杀菌剂交替或混合使用,以延缓抗药性产生和发展。     

Identification of a novel point mutation in the <Emphasis Type="Italic">β</Emphasis>-tubulin gene of <Emphasis Type="Italic">Botrytis cinerea</Emphasis> and detection of benzimidazole resistance by a diagnostic PCR-RFLP assay

The molecular basis of resistance to benzimidazole fungicides with laboratory and field mutant isolates of Botrytis cinerea was investigated. After chemical mutagenesis with N-methyl-N-nitrosogouanidine (NMNG) two different benzimidazole-resistant phenotypes were isolated on media containing carbendazim or a mixture of carbendazim and diethofencarb. The mutant isolates from the fungicide-mixture-containing medium were moderately resistant to carbendazim with wild-type tolerance to diethofencarb while mutant isolates from carbendazim-containing medium were highly resistant to carbendazim but sensitive to diethofencarb. The studied field isolates were highly resistant to benzimidazoles and sensitive to diethofencarb. Study of fitness characteristics of benzimidazole highly-resistant isolates showed that the resistance mutation(s) had no apparent effect on fitness-determining parameters. Contrary to this, the moderately benzimidazole-resistant strains, with no increased diethofencarb sensitivity, had a significant reduction in certain ecological fitness-determining characteristics. Analysis of the sequence of the β-tubulin gene revealed two amino acid replacements in the highly benzimidazole-resistant mutants compared to that of the wild-type parent strain. One was the glutamic acid (GAG) to alanine (GCG) change at position 198 (E198A), identified in both laboratory and field highly benzimidazole-resistant isolates, a mutation previously implicated in benzimidazole resistance. The second was a novel benzimidazole resistance mutation of glutamic acid (GAG) to glycine (GGG) substitution at the same position 198 (E198G), identified in a highly benzimidazole-resistant laboratory mutant strain. Molecular analysis of the moderately benzimidazole-resistant strains revealed no mutations at the β-tubulin gene. A novel diagnostic PCR-RFLP assay utilising a BsaI restriction site present in the benzimidazole-sensitive (E198) but absent in both resistant genotypes (E198G and E198A) was developed for the detection of both amino acid replacements at the β-tubulin gene.     

烟草黑胫病菌抗甲霜灵突变体的诱导及其适合度研究

室内离体条件下采用药剂诱导法,获得了11个烟草黑胫病菌抗甲霜灵突变体;检测了所有抗药突变体的抗药稳定性及抗性水平;比较了3个低抗和1个高抗突变体的适合度。结果表明:在2~20 μg/mL范围内,抗药突变体的发生频率随甲霜灵诱导浓度的增加而提高;所有抗药突变体的抗药性都能稳定遗传;11个抗药突变体中,10个为低抗突变体(抗性水平3.55~13.9倍,平均8.60倍),1个为高抗突变体(抗性水平510倍);低抗菌株20M-001、10M-003、10M-006和高抗菌株10M-004的适合度指数分别为0.56、0.38、0.38和0.87;抗药突变体的适合度指数与抗性水平不相关。     

Characterization of mutations in the iron-sulphur subunit of succinate dehydrogenase correlating with Boscalid resistance in Alternaria alternata from California pistachio

Thirty-eight isolates of Alternaria alternata from pistachio orchards with a history of Pristine (pyraclostrobin + boscalid) applications and displaying high levels of resistance to boscalid fungicide (mean EC(50) values >500 microg/ml) were identified following mycelial growth tests. A cross-resistance study revealed that the same isolates were also resistant to carboxin, a known inhibitor of succinate dehydrogenase (Sdh). To determine the genetic basis of boscalid resistance in A. alternata the entire iron sulphur gene (AaSdhB) was isolated from a fungicide-sensitive isolate. The deduced amino-acid sequence showed high similarity with iron sulphur proteins (Ip) from other organisms. Comparison of AaSdhB full sequences from sensitive and resistant isolates revealed that a highly conserved histidine residue (codon CAC in sensitive isolates) was converted to either tyrosine (codon TAC, type I mutants) or arginine (codon CGC, type II mutants) at position 277. In other fungal species this residue is involved in carboxamide resistance. In this study, 10 and 5 mutants were of type I and type II respectively, while 23 other resistant isolates (type III mutants) had no mutation in the histidine codon. The point mutation detected in type I mutants was used to design a pair of allele-specific polymerase chain reaction (PCR) primers to facilitate rapid detection. A PCR-restriction fragment length polymorphism (RFLP) assay in which amplified gene fragments were digested with AciI was successfully employed for the diagnosis of type II mutants. The relevance of these modifications in A. alternata AaSdhB sequence in conferring boscalid resistance is discussed.     

辣椒疫霉对双炔酰菌胺的敏感性测定及其 抗药突变体生物学性状研究

研究了辣椒疫霉对双炔酰菌胺的敏感性及其抗药性风险。采用菌丝生长速率法测定了分别采自河北定兴、藁城和山东寿光、青州等地的173株辣椒疫霉菌株对双炔酰菌胺的敏感性,结果表明,其EC50值介于0.002 7~0.049 9 μ g/mL之间,平均EC50值为0.021 9±0.010 2 μ g/mL,敏感性呈单峰曲线分布;采用紫外诱导+菌落角变和药剂驯化的方法分别处理10个供试菌株,各获得7株和1株突变体,其抗性倍数在41.50~394 437.77之间,突变频率分别为1.46%和1.43%。对突变体部分生物学性状的研究表明,抗性突变体与其亲本菌株在菌丝生长速率、致病性、产孢能力、适合度指数和竞争力方面均具有不同程度的差异;抗性突变体DX 2-3 20m和DX 2-3 F7的产孢量、适合度和竞争力均高于亲本菌株,有利于抗性群体的形成。研究表明,辣椒疫霉对双炔酰菌胺存在中等抗性风险。     

番茄叶霉病菌对多菌灵抗药性的诱导及抗性菌株特性研究

采用紫外线诱导及药剂驯化两种方法对番茄叶霉病菌的目标菌株进行多菌灵抗性诱导,分别经7、9代诱导后获得抗性突变体。突变体菌株的EC₅₀均大于500μg／mL,达高抗水平。突变体菌株与自然抗性菌株无药继代培养10代后,抗性程度没有明显变化。与亲本菌株比较,突变体菌株菌落生长速率、产孢量及产毒量有所下降。室内交互抗性测定表明:多菌灵与苯菌灵、菌核净及克霉灵之间具有正交互抗性,与瑞毒霉、扑海因及速克灵无交互抗性。乙霉威对多菌灵的高抗菌株表现负交互抗性或无交互抗性,但对敏感菌株不表现负交互抗性。     

番茄早疫病菌对异菌脲的敏感基线及其抗性突变体的生物学特性

采用菌落直径法测定了山西省五寨县42株番茄早疫病菌Alternaria solani菌株对异菌脲的敏感性。结果表明,异菌脲对各菌株的EC50值在1.225~2.674 μ g/mL之间,平均为2.556 μ g/mL, 由于所有菌株均采自未使用过异菌脲的地区,该平均值可作为番茄早疫病菌对异菌脲的相对敏感基线。为了评价番茄早疫病菌对异菌脲的抗性风险,以敏感菌株为试材,通过紫外诱导和药剂驯化(10代)的方法获得了21株抗性突变体,比较了敏感菌株和突变体的生物学特性。结果表明:异菌脲对突变体的EC50值均大于100 μ g/mL,达到高抗性水平;突变体的抗药性稳定,在无药PDA平板上继代培养10次后抗性不丧失;突变体在生长速率及致病力方面较敏感菌株有所下降,产孢量及分生孢子萌发率与敏感菌株相比无显著差异,孢子的竞争能力明显弱于敏感菌株。